Correction: Genetic characterization of extended-spectrum ß-Lactamase- and carbapenemase-producing Escherichia coli isolated from Egyptian hospitals and environments.

[This corrects the article DOI: 10.1371/journal.pone.0255219.].

Effect of Resveratrol and Curcumin on Gene Expression of Methicillin-Resistant Staphylococcus aureus (MRSA) Toxins.

Staphylococcus aureus is an opportunistic pathogen that can lead to a number of potentially terrible community- and hospital-acquired illnesses. Among the diverse set of virulence factors that S. aureus possesses, secreted toxins play a particularly preeminent role in defining its virulence. In this work, we aimed to facilitate the development of novel strategies utilizing natural compounds to lower S. aureus's toxin production and consequently enhance therapeutic approaches. Two natural polyphenols, resveratrol (RSV) and curcumin (CUR), were tested for their effect on reducing toxin gene production of MRSA isolates. Fifty clinical MRSA isolates were gathered from Riyadh and Jeddah. Molecular screening of toxin genes (sea, seb, sec, sed, seh, lukF, and lukS) harbored by MRSA was performed. Sub-inhibitory concentrations of RSV (50 µg/ml) and CUR (20 µg/ml) were determined to study their effect on the gene expression MRSA's toxin genes. Our findings revealed the presence of the tested genes in MRSA isolates, with lukF being the most prevalent gene and seh the least detected gene. We found that RSV reduced the relative expression of toxin genes, sea, seb, lukF, and lukS, respectively, while CUR decreased the relative expression of sea and seb genes in the examined isolates. Regarding lukF and lukS, CUR downregulated the expression of both genes in some isolates and upregulated the expression in other isolates. From these results, we concluded that RSV and CUR could be used as alternative therapeutic approaches to treat MRSA infections through reducing toxin production.

Characterization of Integrons and Quinolone Resistance in Clinical Escherichia coli Isolates in Mansoura City, Egypt.

Escherichia coli is a common pathogen in both humans and animals. Quinolones are used to treat infections caused by Gram-negative bacteria, but resistance genes emerged. Only scarce studies investigated the association between plasmid-mediated quinolone resistance (PMQR) genes and integrons in clinical isolates of E. coli. The current study investigated the prevalence of quinolone resistance and integrons among 134 clinical E. coli isolates. Eighty (59.70%) isolates were quinolone-resistant, and 60/134 (44.77%) isolates were integron positive with the predominance of class I integrons (98.33%). There was a significant association between quinolone resistance and the presence of integrons (P < 0.0001). Isolates from Urology and Nephrology Center and Gastroenterology Hospital were significantly quinolone-resistant and integron positive (P ≤ 0.0005). Detection of PMQR genes on plasmids of integron-positive isolates showed that the active efflux pump genes oqxAB and qepA had the highest prevalence (72.22%), followed by the aminoglycoside acetyltransferase gene (aac(6')-Ib-cr, 66.67%) and the quinolone resistance genes (qnr, 61.11%). Amplification and sequencing of integrons' variable regions illustrated that no quinolone resistance genes were detected, and the most predominant gene cassettes were for trimethoprim and aminoglycoside resistance including dfrA17, dfrB4, and dfrA17-aadA5. In conclusion, this study reported the high prevalence of PMQR genes and integrons among clinical E. coli isolates. Although PMQR genes are not cassette-born, they were associated with integrons' presence, which contributes to the widespread of quinolone resistance in Egypt.

Genetic characterization of extended-spectrum ß-Lactamase- and carbapenemase-producing Escherichia coli isolated from Egyptian hospitals and environments.

Over the past decades, Escherichia coli (E. coli) have acquired extensive resistance to antibiotics; especially ß- lactams. This study aimed to investigate the frequency of Extended-spectrum ß-lactamase (ESBL) and carbapenemase producers among E. coli isolates and their correlation with serotypes, phylogenetic background, and pathogenicity associated islands. A total of 105 E. coli strains were isolated and subjected to antimicrobial susceptibility testing against ß-lactam antibiotics. All isolates showed a high resistance profile. Resistant isolates were tested for ESBL and carbapenemase production. Fifty-three and 18 isolates were positive for ESBL and carbapenemase producers, respectively. ESBL and carbapenemase genes were detected by PCR. TEM gene was the most prevalent gene among all isolates followed by SHV and CTX-M15. In carbapenemase-producers, OXA-48 and IMP were the predominant genes. Enteropathogenic E. coli (EPEC) and Enterohemorrhagic E. coli (EHEC) were the major producers of ESBL and carbapenemase, respectively as indicated by serodiagnosis. They were further assessed for the presence of pathogenicity islands (PAIs) and phylogenetic background. The most predominant DEC PAI and ExPEC PAI were HPI and IICFT073. Most clinically ESBL-producers were group D and B2 while environmentally ones were group B1 and A. On contrary, clinically carbapenemase-producers belonged to group C and D. In conclusion, our study confirms the importance of phylogenetic group D, B2, and C origin for antibiotic resistance in E. coli. Ultimately, our findings support the fact that environmental isolates contribute to the local spread of E. coli pathogenicity in Egypt and these isolates maybe serve as reservoirs for transmission of resistance.

Comparative Assessment of Different PCR-Based Typing Methods of Pseudomonas aeruginosa Isolates.

INTRODUCTION: Pseudomonas aeruginosa is one of the important causes of nosocomial infections. Analyzing the diversity of these isolates is important to control the diseases caused by them. Studies of molecular epidemiology depend on the application of typing methods. PURPOSE: This study aims to assess the performance of PCR- based typing techniques (RAPD, ribotyping, tDNA, and ERIC) in determining the genetic diversity of 44 P. aeruginosa urinary isolates. METHODS: Performance parameters were analyzed for each of the tested methods. The banding pattern was assessed by calculating polymorphism, genotypic gene diversity and the effective multiplex ratio. Moreover, strain diversity, typeability, and discriminatory power were used to measure the efficiency of typing methods. The congruence among typing methods was calculated by Rand's and Wallace coefficients. RESULTS: P-640 among RAPD primers and Ribo-2 among ribotyping primers were more informative as they gave high strain diversity, the highest number of clusters, and highest discriminatory power (ISD=70.45%, 29 clusters at 70% cutoff, DI=0.97 and ISD=75%, 25 clusters at 70% cutoff DI=0.969, respectively). Comparison of typing methods showed that RAPD-PCR gave the highest mean percent polymorphism per assay (76.85%) followed by ERIC-PCR. ERIC-PCR outperformed in most marker parameters; highest mean number of alleles, number of monomorphic bands per assay unit, mean genotypic gene diversity, effective multiplex ratio, and assay efficiency index. Calculated congruence revealed that individual methods demonstrate moderate to poor predictive power. Interestingly, this power increased by combining data obtained from another method. CONCLUSION: RAPD primer (P-640) had more discrimination power followed by ribo-2 and ERIC. The performance and predictive power of typing methods can be improved by combining data obtained from different methods as ERIC+OPA-02 and ERIC+P-640 combinations gave complete typeability and discrimination of isolates. ERIC, ERIC+OPA-02, and ERIC+P-640 combinations can provide finer discrimination and classification of P. aeruginosa strains than the other tested methods.

Effect of Sub-Minimum Inhibitory Concentrations of Tyrosol and EDTA on Quorum Sensing and Virulence of Pseudomonas aeruginosa.

INTRODUCTION: Pseudomonas aeruginosa is considered a dangerous pathogen, as it causes many human diseases, besides that it is resistant to almost all types of antibacterial agents. So, new strategies to overcome P. aeruginosa infection have evolved to attenuate its virulence factors and inhibit its quorum-sensing (QS) activity. PURPOSE: This study investigated the effect of tyrosol and EDTA as anti-quorum-sensing and antivirulence agents against P. aeruginosa PAO1. METHODS: Anti-quorum activity of sub-minimum inhibitory concentrations (sub-MICs) of tyrosol and EDTA was tested using Chromobacterium violaceum (CV 12,472) biosensor bioassay. Miller assay was used to assess the inhibition of QS signal molecules by ß-galactosidase activity determination. Also, their effects on the production of protease, lipase, lecithinase, and motility were tested. The inhibitory effects of these molecules on QS regulatory genes and exotoxins genes expression were evaluated by real-time PCR. RESULTS: Tyrosol and EDTA at sub-MICs inhibited the production of violacein pigment. Both compounds inhibited QS molecules production and their associated virulence factors (protease, lipase, lecithinase, and motility) (P≤ 0.05). Besides, the expression levels of QS regulatory genes (lasI, lasR, rhƖI, rhIR, pqsA, and pqsR) and exotoxins genes (exoS and exoY) were significantly reduced (P≤ 0.05). CONCLUSION: Both tyrosol and EDTA can be used to fight P. aeruginosa infection as anti-quorum-sensing and antivirulence agents at their sub-MICs.

Characterization of ß-lactam resistance in K. pneumoniae associated with ready-to-eat processed meat in Egypt.

K. pneumoniae was known as a nosocomial infection that causes human diseases. It is considered as one of the food-borne pathogens as it causes septicemia and diarrhea in humans. This study aims to characterize K. pneumoniae strains isolated from ready to eat processed meat phenotypically and genetically. Three hundred and fifty ready to eat processed meat (Luncheon-meat) samples were collected. Forty-four (12.6%) K. pneumoniae strains were isolated and bio-typed, where the majority were identified to belong to biotype B1. K1 and K2 serotypes were detected and strains were classified as hypermucoviscous K. pneumoniae (HVKP) and classic K. pneumoniae (CKP) (26 and 18 isolates, respectively). The isolates were resistant to several classes of ß-lactam antibiotics, ceftazidim and cefotaxime (95.5%), cefoxitin (93.2%), ertapenem (90.9%) and amoxicillin-clavulanic acid (86.4%). They were classified as extended spectrum ß-lactamases (ESBLs), AmpC or carbapenemase-producers phenotypically. Eighteen ß-lactamase genes were investigated by PCR. The most prominent genes were SHV (63.6%), TEM (52.2%), CTX-M15 (50%), AMPC (47.7%), CIT-M (45.5%) and VIM (43.2%). Co-detection of ß-lactam resistance genes revealed 42 gene profiles. Twenty-four isolates had the complete efflux system (AcrAB-ToƖC). Besides, Integrons (I, II, III) were detected in 20 isolates. Molecular typing by ERIC-PCR showed high genetic diversity between isolates as 34 different patterns were identified. Overall, this study confirmed the hazards posed by the presence of multiple resistance genes in the same isolate and this should not be undervalued. Besides, the horizontal transfer of plasmid harboring resistance genes between isolates in food represents potential health risks for consumers in Egypt and so the control and inhibition plans are necessary.